Elucidating molecular mechanisms that are altered during HIV-1 infection may provide a better understanding of the HIV-1 life cycle and how it interacts with infected T-cells. One such mechanism is alternative splicing (AS), which has been studied for HIV-1 itself, but no systematic analysis has yet been performed on infected T-cells. We hypothesized that AS patterns in infected T-cells may illuminate the molecular mechanisms underlying HIV-1 infection and identify candidate molecular markers for specifically targeting infected T-cells. We downloaded previously published raw RNA-seq data obtained from HIV-1 infected and non-infected T-cells. We estimated percent spliced in (PSI) levels for each AS exon, then identified differential AS events in the infected cells (FDR < 0.05, PSI difference > 0.1). We performed functional gene set enrichment analysis on the genes with differentially expressed AS exons to identify their functional roles. In addition, we used RT-PCR to validate differential alternative splicing events in cyclin T1 (CCNT1) as a case study. We identified 427 candidate genes with differentially expressed AS exons in infected T-cells, including 20 genes related to cell surface, 35 to kinases, and 121 to immune-related genes. In addition, protein-protein interaction analysis identified six essential subnetworks related to the viral life cycle, including Transcriptional regulation by TP53, Class I MHC mediated antigen, G2/M transition, and late phase of HIV life cycle. CCNT1 exon 7 was more frequently skipped in infected T-cells, leading to loss of the key Cyclin_N motif and affecting HIV-1 transcriptional elongation. Our findings may provide new insight into systemic host AS regulation under HIV-1 infection and may provide useful initial candidates for the discovery of new markers for specifically targeting infected T-cells.

中文翻译：

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HIV-1感染的CD4 T细胞中选择性剪接的情况

阐明在HIV-1感染过程中发生改变的分子机制可以更好地了解HIV-1的生命周期以及它如何与感染的T细胞相互作用。这样的机制之一是替代剪接（AS），该替代剪接已针对HIV-1本身进行了研究，但尚未对感染的T细胞进行系统的分析。我们假设感染的T细胞中的AS模式可能阐明了HIV-1感染的分子机制，并确定了专门针对感染T细胞的候选分子标记。我们下载了先前发布的从HIV-1感染和未感染T细胞获得的原始RNA-seq数据。我们估算了每个AS外显子的剪接百分比（PSI），然后确定了感染细胞中的AS差异事件（FDR <0.05，PSI差异> 0.1）。我们对差异表达的AS外显子的基因进行了功能基因集富集分析，以鉴定其功能作用。此外，作为案例研究，我们使用RT-PCR验证了细胞周期蛋白T1（CCNT1）中不同的可变剪接事件。我们在被感染的T细胞中鉴定了427个候选基因，它们具有差异表达的AS外显子，包括20个与细胞表面相关的基因，35个与激酶相关的基因和121个与免疫相关基因的基因。此外，蛋白质-蛋白质相互作用分析确定了与病毒生命周期相关的六个基本子网，包括TP53的转录调控，I类MHC介导的抗原，G2 / M过渡和HIV生命周期的后期。CCNT1外显子7在感染的T细胞中更常被跳过，导致关键Cyclin_N基序丢失并影响HIV-1转录延伸。